laboratory handling of the renal biopsy dr. issam francis kuwait 4 th ssn annual international...
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Laboratory Handling of the Renal BiopsyDr. Issam Francis
Kuwait
4th SSN Annual International Conference, Riyadh, April 2009
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The Renal Biopsy Historical backgrounds
• 1934 Percutaneous kidney biopsy (tumors) • 1951 first kidney biopsy for medical disease • 1953 Introduced to US• 1955 first renal clinicopathology working group
Modern Times:• Real-time ultrasound guidance• Transjugular needle biopsy• Spring-loaded biopsy gun• CT-Guided biopsy• EM• IHC
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Needle Biopsy Open Biopsy
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Open Biopsy
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The Renal Biopsy Laboratory handling
EM Paraffin Sections IF
Under a stereomicroscope
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The Renal Biopsy Laboratory handling
EM Paraffin Sections & IHC (no IF)
Under a stereomicroscope
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The Renal Biopsy Laboratory handling
1- fixation (Immediate):• 10 % NB Formalin (paraffin sections)• 4% Gluteraldehyde (EM)• No fixation (Immunofluorescence)
2- Paraffin sections cut at 3 u thickness
3- Stains: HE PAS GMS TC CR …..HE PAS GMS HE TC HE PAS GMS HETC HE HE PAS GMS HE TC PAS
GMSHE TC (CR, Microbial stains, others.)
4- Immunohistochemistry (IG, C, other antigens)
5- Immunofluorescence (IG, C)
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The Renal Biopsy Laboratory handling
• HE• PAS• JNS• TC• HE• PAS• JNS• TC
3-4 microns24 Sections
Extra sectons for CR, Microbial stains, IHC etc.
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The Renal Biopsy Laboratory handling
Biopsy adequacy:– Cortex and medulla– 1-2 glomeruli EM– 3-5 glomeruli IF– 6 glomeruli PS (native kidney)– 10 glomeruli PS (renal allograft)
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The Renal Biopsy Manual vs. automated
05
1015202530354045
Man
ual
Au
tom
ated
# of cores
Totalglomeruli
Glomeruliper core
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Biopsy Fixation
10% Neutral Buffered Formalin Solution:Why?1. Cheap2. Commonly available3. Suitable for:
– All histological stains– Immunohistological methods (not IF)
4. Reversible: Possible to transfer to another fixative for electron microscopy.
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Morphological examination:1. Glomeruli2. Tubules3. Interstitium4. Blood vessels
Renal Biopsy Morphological Examination
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Primary Site of Renal Pathology
Glomerulus• Glomerular pathology:
– Inflammation– GBM changes– Scarring– Abnormal deposits– Cellularity
Tubules • Tubular pathology :
– Cellular injury– Regeneration – Atrophy – Casts
Interstitium • Interstitial
pathology:– Cellular infiltrates– Edema/fibrosis
Vascular disease • Vascular pathology:
– Inflammation– Sclerosis– Hyalinosis– Thrombosis
? Stain
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Activity of disease:• Cellular proliferation• Crescent formation• Necrotizing lesions• Inflammation
Chronicity of disease :• Tubular atrophy• Fibrosis• Vascular sclerosis
Renal pathology report Disease Stage
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The Renal Biopsy Stains1. HE General
2. PAS Basement M. & Mesangial matrix
3. Trichrome Fibrosis
4. Silver Basement M. & Mesangial matrix
5. Congo red Amyloid
6. MSB Fibrin
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Masson’s trichrome
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Structure/Component PAS Jones Masson’s trichrome
Basement membrane Red Black Deep blue
Mesangial matrix Red Black Deep blue
Interstitial collagen Negative Negative Pale blue
Cell cytoplasm (normal) Negative (most) Negative Rust/orange
Immune complex Negative Negative Bright red
Amyloid Negative Negative Light blue
Tubular casts Red Gray to black Light blue
Staining characteristics of selected normal and abnormal renal structures
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SATINING OF RENAL TISSUE COMPONENTS
FEATURE HE PAS TRICHROME JONES/GMS
Cellularity Excellent Excellent Poor Poor
Mesangial M Poor Excellent Variable Excellent
Glom. Sclerosis Poor Excellent Excellent Good
Immune Cox. Poor Poor Variable Negative
Basement M. Poor Excellent Good Excellent
Fibrosis Poor Poor Excellent Excellent
Vascular hyaline Good Poor Good Negative
Thrombi Good Poor Good Variable
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H&E
Amyloid:• Silver negative• 8 micron sections
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Congo Red
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Fibrillary GP
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Diagnoses overlooked without IHC
1. Light chain-associated diseases2. AL amyloid3. Monoclonal immunoglobulin deposition disease4. Light chain cast nephropathy5. IgA nephropathy/Henoch–Shonlein purpura6. IgM nephropathy7. C1q nephropathy8. Antiglomerular basement membrane disease9. Humoral (C4d) transplant rejection10. Fibronectin glomerulopathy
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IgG
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IgA
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Electron microscopic helpful, but not usually essential for the diagnosis1. Nephrotic syndrome2. Acute renal failure3. Chronic renal failure4. Renal disease in diabetes mellitus5. Renal disease in SLE6. Suspected rejection of a renal allograft7. Repeat specimen when the diagnosis has
been made
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Electron microscopy is most likely to help
1. Microscopic Hematuria with normal renal function
2. Family history of renal disease.3. Asymptomatic proteinuria with normal renal
function.
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Diagnoses overlooked without EM
1. Fibrillary /immunotactoid glomerulopathy2. Nail–patella syndrome3. Lipoprotein glomerulopathy4. Dense deposit disease5. Alport’s syndrome6. Thin basement membrane nephropathy7. Collagenofibrotic glomerulopathy
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Thank You